1. Field of the Invention
The present invention concerns a method of enzymatically determining the pH of a specimen (e.g., a solution or a biological fluid) and a kit for conducting the method.
2. Discussion of the Background
Enzyme activities are generally known to be pH dependent. Typically, a graph showing the relationship of enzymatic activity (plotted along the ordinate) to pH (plotted along the abscissa) contains a bell-shaped curve (e.g., FIG. 1). Although such information is useful for predicting enzymatic activity as a function of pH (e.g., for determining optimum pH and maximizing enzymatic activity), the converse relationship does not necessarily exist; that is, one cannot conclusively determine the pH of a sample containing the enzyme from the enzyme activity information alone. A horizontal line corresponding to a given enzymatic activity intersects a bell-shaped curve such as that in FIG. 1 at two or more points. As a result, from a given set of test conditions, more than one pH value may correspond to a given enzymatic activity.
Drug testing is becoming an increasingly important tool for improving safety, security and productivity in a broad range of industries and commercial activities (e.g., transportation, medicine, defense, government, etc.). A number of Federal guidelines exist for implementing and conducting drug testing (see, for example, Executive Order No. 12564, "Mandatory Guidelines for Federal Workplace Drug Testing Programs," Federal Register, September 15, 1988).
The integrity and proper identity of specimens collected are critical for success in drug testing. For example, a urine-based drug testing program normally involves the steps of (1) specimen collection, (2) an initial-immunoassay based screening test (3) confirmation testing based on gas chromatography/mass spectrometry (GC/MS), and (4) the distribution of test results. Many drug users attempt to evade drug detection by adulterating the specimen. Such adulteration is intended to produce a false negative test result during initial immunoassay screening.
Common adulteration methods include dilution with water, substitution of a drug-free liquid, and addition of household materials or chemicals (e.g., vinegar, baking soda, table salt, lye [e.g., DRANO.RTM.], detergent, or a substance containing glutaraldehyde such as URINE-AID, etc.). A drug user may also attempt alter his or her urinary pH (i.e., urinary acidity or alkalinity) to facilitate faster elimination of certain drugs or drug metabolites through metabolic processes. This latter technique may be applicable to elimination of alkaloids, amphetamines and phencyclidine, for example.
A number of methods may detect or deter urine adulteration, including temperature measurement, direct urine pH measurement, determination of the specific gravity of the sample and/or determination of the presence and/or concentration of creatinine. "Normal" urine should have a temperature of 32.5.degree.-37.7.degree. C. (90.5-99.8.degree. F.), a pH of 5-8, a specific gravity of 1.003-1.030 and a creatinine concentration of 0.8-1.4 mg/dL. If any of these parameters are outside the specified ranges, one has reason to suspect the urine specimen has been adulterated.
Prior to the present invention, urine pH measurement has been determined by use of pH paper or by an endpoint colorimetric pH measurement method (e.g., the pH PERFECT.RTM. drug testing adulteration testing kit, available from Chimera Research & Chemical, Inc., Seminole, Fla.). However, methods using pH paper are slow, subjective and cannot be adapted to existing clinical chemistry analyzers for high-volume urine screening applications. Colorimetric methods typically suffer from a lack of an indicator which is suitable for applications over a broad pH range (i.e., pH 2-12).
As a result, a need exists for a simpler, more accurate method of determining the pH of a specimen (e.g., a solution or a biological fluid) which may be more applicable for high volume screening (e.g., urine testing), a more direct method of testing an adulterated specimen through pH measurement, and for a test kit for conducting such methods.